Regenerative medicine is aimed to regenerate the architecture and function of tissues and organs totally or partially lost due to disease, trauma and ageing (Vats et al., Lancet 366:562-602, 2005). Stem cells are considered crucial building blocks for any regenerative strategy. The challenge and motivation are to find the ways for recruiting and/or delivering to the injured site pluripotent stem cells populations capable of regenerating nonfunctional or lost tissues and organs. The restricted location of tissues resident stem cells and the relative low number of circulating stem cells pose serious doubts whether in vivo recruitment of endogenous stem cells may be a realistic clinical prospect.
There are currently three possible strategies for regenerative stem cell-based therapies: a) stimulation of resident stem cells of the affected tissue. Resident stem cells (SC) have been identified in numerous tissues including in those that are most frequently affected by disease as cardiac, neural, pancreatic, and kidney tissues; b) recruitment of circulating bone marrow-derived multipotent stem cells to the site of damage. These have been identified and found to increase in number following acute damage; c) transplantation of ex-vivo expanded stem cells. In this respect, the challenge is to generate a pluripotent population of stem cells that can be expanded ex-vivo without losing its potency and can be safely transplanted with no adverse reactions.
Three major optional sources for stem cells are currently being investigated i) Embryonic stem cells (ECS); ii) Fetal stem cells (FSC); and iii) Adult (somatic) stem cells (ASC) either autologous or allogeneic.
ESC lines are pluripotent and virtually immortal (Thomson et al., Science 282:1145-1147, 1998), but are still far from clinical applications because of their tendency to undergo transformation when implanted in vivo, their antigenicity (Bradley et al., Nat Rev Immunol 2:859-871, 2002; Drucker et al., Trends in Biotechnology. 22:136-141, 2004), the requirement for a mouse feeder layer for their maintenance and expansion and ethical issues.
Theoretically, FSC could be ideal for therapy as on the one hand they might be sufficiently differentiated not to undergo transformation following implantation and on the other hand they still retain sufficient pluripotent properties. Because of ethical reasons only FSC derived from the umbilical cord are currently utilized. However, similarly to allogeneic ASC, umbilical FSC are considered allogeneic transplantations and therefore their use requires antigenic matching, immunosuppressive therapy and can result in rejection or graft versus host disease (Brunstein et al., Vox Sanguinis 91:195-205, 2006).
Thus, an autologous pluripotent stem cells population derived from the adult is considered the ideal population for tissue and organ regeneration. Bone marrow and to very limited extent, peripheral blood, adipose tissue, skin and muscle are the major sources for autologous adult stem cells. A serious drawback of these sources is that aging and disease substantially lower the functionality and possibly the availability of adult stem cells in the bone marrow and other tissues (Janzen et al., Nature 28; 443 (7110):421-26, 2006; Rando T A. Nature 441:1080-1086, 2006; Sethe et al., Aging Res Rev 5:91-116, 2006. Rao and Mattson, 122:713-734, 2001). Another considerable drawback of bone marrow-derived pluripotent stem cells is their rarity and cumbersome isolation procedures (Reyes M et al., Blood 96:2615-2625, 2001; D'Ippolito G et al., Rejuvenation Research 9:10-18, 2006).
Aging and associated diseases decrease tissue response to stress. This has been in part attributed to reduction in the tissue specific stem cells functionality as reflected by reduced self-renewal, homing and engraftment abilities (Janzen et al., Nature 28; 443 (7110):421-26, 2006).
Although the reasons for these changes are still debated, they are considered inherent to aging and/or imposed by epigenetic factors such as changes in stem cells niche.
US 2005/0032207 discloses a method for isolating, culturing and differentiating intestinal stem cells for therapeutic uses. According to this application undifferentiated somatic intestinal stem cells are isolated from intestinal epithelium of a tissue which can comprise portion of the: stomach, duodenum, jejunum, ileum, cecum, colon, e.g. ascending colon, transverse colon, descending colon, sigmoid colon, rectum, anal canal, and/or appendix.
Oral Mucosa
Oral Mucosa is the mucosal lining the oral cavity (FIG. 1). It is a complex tissue consisting of cell populations and the extracellular matrix which houses the cells and provides a substrate for cell attachment. Oral mucosa consists of an epithelial tissue of ectodermal origin and the lamina propria which is a connective tissue of ectomesenchymal origin (FIG. 2). The epithelial part comes into contact with the oral cavity and functions as a barrier that is continuously renewed by a unipotent stem cell population located on the basement membrane, a structure which connects the epithelial and ectomesenchymal parts together. The lamina propria supports the epithelial tissue and is attached to the underlying structures of the oral cavity which are protected by the oral mucosa. Clinical observations indicate that full thickness incisional and excisional surgical wounds in the oral mucosa heal substantially faster than wounds in other connective tissues (Cate RA. Repair and Regeneration of Oral Tissues in Ten Cate's Oral Histology sixth edition 2003; Stephens et al., J Dent Res. 75 (6):1358-1364, 1996) and that similarly to early fetal mammalian tissues and a number of adult tissues in low vertebrates, wounds in the oral mucosa heal by regeneration and not by scar formation (Hallock G G., Plastic Reconstructive Surgery 75:785-788, 1985).
It has been disclosed that stem cells suitable for use in therapy may be obtained from various sources. Laino et al. (J. Cell. Physiol. 206, 693-701, 2005) discloses an approachable human adult stem cell source derived from dental pulp (i.e., the cell mass within the tooth), particularly suitable for hard-tissue engineering. U.S. Pat. No. 7,052,907 discloses and claims a culture of isolated human dental pulp stem cells. US Patent Application Publication No. 2006/0252151 discloses methods for harvesting stem cells from dental pulp.
U.S. Pat. No. 7,015,037 discloses the isolation of multipotent adult progenitor cells derived from bone marrow, characterization and uses thereof. In particular the multipotent adult progenitor cells co-express CD 49c and CD 90 and have a doubling time of about 36 hours.
U.S. Pat. No. 7,078,230 discloses pluripotent stem cells generated from adipose tissue-derived stromal cells and uses thereof. In particular, the invention includes isolated adipose tissue derived stromal cells that have been induced to express at least one phenotypic characteristic of a neuronal, astroglial, hematopoietic progenitor, or hepatic cells.
PCT publication WO 2004/009758 discloses a method for isolating embryonic-like stem cells from adult sources using the germ stem cell markers expressed in primordial stem cells but not in differentiated somatic cell types.
PCT publication WO 2003/089631 discloses a method for propagating stem cells and/or progenitor cells which may be used with isolated adult human tissue optionally obtainable from the olfactory lamina propria. PCT publication WO 2007/020611 discloses adult human neural stem cells from the olfactory lamina propria.
Thus, there is still an unmet need in the art to identify a readily accessible source of stem cells in mammals, capable of generating an autologous pluripotent population of stem cells that can be expanded in vitro without losing its pluripotency and can be safely retransplanted into the affected donor to achieve tissue and organ regeneration effectively.